1. Field of the Invention
The present invention relates to the method of mass analysis using the time-of-flight (TOF) mass spectrometry and, more specifically, it relates to TOF mass spectrometry in which a reflectron is used for correction of the initial ion velocity distribution in an ion source, and, most specifically, it relates to the method of designing a reflectron for a TOF mass spectrometer which takes into account the effect of presence of acceleration and deceleration regions in the ion source, ion detector, and other ion optics elements of the TOF mass spectrometer on the correction of the initial ion velocity distribution. The invention also relates to an improved mass spectrometer design and, more specifically, it relates to such a design for achieving high mass resolution in the matrix-assisted laser desorption/ionization (MALDI) TOF mass spectrometer, and, most specifically, it relates to the design of a high resolution MALDI/TOF mass spectrometer using low voltages for extraction of ions from an ion source.
2. Description of the Related Art
The time-of-flight (TOF) mass spectrometer (MS) is a well known instrument for mass analysis in which ions formed from sample molecules in an ion source are accelerated to the same energy and allowed to drift along some path before detection. Because ions of different mass have different velocity after acceleration they are separated in space during flight and in time during detection, thus, the time of arrival to the detector is measure of mass (or mass-to-charge ratio m/z if ions are not singly-charged). However, such a simple picture is always complicated by the presence of non-ideal factors and among them are: (a) different time of formation or acceleration of ions; (b) different initial location of ions in space; and (c) different initial velocity of ions before acceleration, as described in Cotter, R. J. Time-of-Flight Mass Spectrometry: Instrumentation and Applications in Biological Research, ACS Professional Reference Books: Washington, 1997; p. 326. However, time focusing can be achieved by using pulsed drawout fields with sharp rise times or short laser pulses in the case of laser desorption (LD) or matrix-assisted laser desorption/ionization (MALDI). A dual-stage extraction method is normally used for correction of the initial spatial distribution of ions in an ion source, as described in Wiley, W. C.; McLaren, I. H. Rev. Sci. Instr. 1955, 26, 1150-1157. And finally, initial velocity (or energy) distribution can be corrected by time-lag focusing technique, which is now also referred to as a pulsed or delayed or time-delayed extraction method. See, for example, Wiley, W. C.; McLaren, I. H. Rev. Sci. Instr. 1955, 26, 1150-1157; Van Breemen, R. B.; Snow, M.; Cotter, R. J. Int. J. Mass Spectrom. Ion Phys. 1983, 49, 35-50; Colby, S. M.; King, T. B.; Reilly, J. P. Rapid Commun. Mass Spectrom. 1994, 8, 865-868; Brown, R. S.; Lennon, J. J. Anal. Chem. 1995, 67, 1998-2003 and Vestal, M. L.; Juhasz, P.; Martin, S. A. Rapid Commun. Mass Spectrom. 1995, 9, 1044-1050.
In the method of mass analyzing and the design of the mass spectrometer which constitute the subject of this patent, the compensation for the effect of initial velocity distribution of ions is of primary concern, which concern is primarily driven by the wide use of MALDI and LD-TOF instruments. Since MALDI and LD ions are desorbed from or formed near a well defined smoothed equipotential surface, the initial spatial distribution of ions is minimized. Initial temporal distribution for ions is also very small due to the use of short pulse lasers (the pulse width of a nitrogen laser is usually less than 1 ns). Thus, in the case of MALDI and LD broadening of the mass spectral lines by the initial velocity distribution is of primary concern. MALDI ions, for example, are desorbed with mean velocities up to hundreds of meters per second, which depend primarily on the matrix, and the energy of desorbed ions may easily reach 10-100 eV depending on ion mass. See, for example, Spengler, B.; Cotter, R. J. Anal. Chem. 1990, 62, 793-796; Ens, W.; Mao, Y.; Mayer, F.; Standing, K. G. Rapid Commun. Mass Spectrom. 1991, 5, 117-123; Huth-Fehre, T.; Becker, C. H. Rapid Commun. Mass Spectrom. 1991, 5, 378-382; Beavis, R. C.; Chait, B. T. Chem. Phys. Lett. 1991, 181, 479-484; Pan, Y.; Cotter, R. J. Org. Mass Spectrom. 1992, 27, 3-8; Zhou, J.; and Ens, W.; Standing, K. G.; Verentchikov, A. Rapid Commun. Mass Spectrom. 1992, 6, 671-678.
The major drawback of the time-delayed extraction method used for the initial velocity distribution correction is its mass dependence, which is very impractical for a TOF mass spectrometer recording the entire mass range. A number of investigators are working on improvements of the time-delayed extraction technique and other methods using dynamic electric fields, such as Marable, N. L.; Sanzone, G. Int. J. Mass Spectrom. Ion Phys. 1974, 13, 185-194; Browder, J. A.; Miller, R. L.; Thomas, W. A.; Sanzone, G. Int. J. Mass Spectrom. Ion Phys. 1981, 37, 99-108; Muga, M. L. Anal. Instrum. 1987, 16, 31-50; Yefchak, G. E.; Enke, C. G.; Holland, J. F. Int. J. Mass Spectrom. Ion Processes 1987, 87, 313-330; Kinsel, G. R.; Johnston, M. V. Int. J. Mass Spectrom. Ion Processes 1989, 91, 157-176; Kinsel, G. R.; Grundwuermer, J. M.; Grotemeyer, J. J. Am. Soc. Mass Spectrom. 1993, 4, 2-10; Kovtoun, S. V. Rapid Commun. Mass Spectrom. 1997, 11, 433-436; and Kovtoun, S. V. Rapid Commun. Mass Spectrom. 1997, 11, 810-815. The problem of mass dependence in the case of using pulsed extraction fields, however, is not yet fully resolved.
Alternatively, an approach to mass independent correction of the initial velocity distribution of ions has been possible with the invention of the ion mirror or the reflectron, described in Mamyrin, B. A.; Karataev, V. I.; Shmikk, D. V.; Zagulin, V. A. Sov. Phys. JETP 1973, 37, 45-48. A reflectron does not actually make any correction of the initial spatial, temporal or velocity distributions but effectively extends the ion flight path, thus, increasing the mass resolution. It simply transfers the temporal and spatial distributions at the start (or focal) plane to the final focal plane formed after reflecting the ions by the ion mirror. As for the initial velocity distribution it is also transferred by the reflectron from the start space focal plane to the final focal plane but with some distortion. The accuracy of the velocity distribution transfer by the reflectron is usually expressed by the order of focusing which is actually the power of the highest zero term in the expansion of the ion time-of-flight over the initial ion velocity. For example, a single-stage linear reflectron performs the first order velocity focusing while a two-stage linear reflectron can focus with the second order accuracy. A parabol ic mirror can perform infinite order focusing, i.e. the ion time-of-flight does not depend on the initial kinetic energy of ions at all, as described in U.S. Pat. No. 4,625,112 to Yoshida. Such mirrors are also referred to as ideal reflectrons. A field inside a parabolic reflectron is curved and according to the LaPlace equation it also has a curvature in a radial (or transverse) direction. This, of course, results in limitations for an angular aperture of such reflectron. Small divergent properties of a parabolic reflectron can also affect the kinetic energy of ions and, thus, the mass resolution. Another feature of a parabolic reflectron, which does not encourage its wide use, is the absence of a field-free region which is always required in TOF/MS for mounting detectors, lenses, energy filters, etc. This drawback is overcome in another design for an ideal reflectron in which a curved field is also used inside the reflectron but the field-free path can be made of any length in comparison with the reflectron length if the minimum initial energy of ions to be focused is allowed to be larger than zero (a parabolic reflectron focuses ions of all energies starting from zero), described by Managadze, G. G.; Shutyaev, I. Yu. In Laser Ionization Mass Spectrometry; Vertes, A.; Gijbels, R.; Adams, F., Eds.; John Wiley and Sons: New York, 1993; p. 505-549. With that design, the most general solution for the field inside an ideal reflectron was obtained. The solutions for some special cases have also been reported using analytical and numerical approaches, described, respectively, in Flory, C. A.; Taber, R. C.; Yefchak, G. E. Int. J. Mass Spectrom. Ion Processes 1996, 152, 177-184 and Vlasak, P.; Beussman, D. J.; Ji, Q.; Enke, C. G. J. Am. Soc. Mass Spectrom. 1996, 7, 1002-1008. A curved-field of special design has been also used in the second reflectron of a tandem MALDI/TOF/TOF instrument for focusing product ions having different kinetic energy after the fragmentation of precursor ions in collisions with target neutral molecules, as described in U.S. Pat. No. 5,464,985 to Cornish, T. J. and Cotter, R. J.
Thus, with the introduction of a reflectron the problem of velocity focusing is reduced to obtaining good conditions at the start focal plane. This is not a problem if the initial spatial distribution in the ion source is the major source of the line broadening in mass spectra because any single or double-stage extraction scheme effectively eliminates the ion space distribution (converting it into the larger velocity distribution of ions at the focal plane). In the case of MALDI where the major contribution into the line broadening comes from the initial velocity distribution of ions the situation is not so clear. It has been shown that velocity focusing cannot be achieved at all in a MALDI/TOF-MS with a single-stage linear reflectron, such as illustrated in FIG. 1A, and the corresponding kinetic energy distribution of injected ions being shown in FIG. 1B. As illustrated and is known, such a single stage reflectron contains an ion source 12, a drawout assembly 14, a reflectron 16 that contains a grid 18, and a detector 20. The same is true when a two-stage linear reflectron is used, such as illustrated in FIG. 2A, which contains essentially the same physical components as the single stage reflectron of FIG. 1A, with a difference being that the reflectron 16 contains two stages, 16A and 16B, to which are applied different voltages V1 and V2 that cause ions to lose their kinetic energy according to the distribution illustrated by FIG. 2B. The use of very high acceleration voltages facilitates but does not solve the problem completely. The problem of velocity focusing in a TOF/MS can be partially solved with specially designed double-stage reflectron 16, such as illustrated in FIG. 3A and the corresponding distribution illustrated in FIG. 3B, or three-stage linear reflectrons that includes stages 16A, 16B and 16C to which are applied voltages V1, V2 and V3, such as illustrated in FIG. 4A and the corresponding distribution illustrated in FIG. 4B, but the accuracy of the velocity focusing is limited by the first and second order correspondingly, as described by Short, R. T., Todd, P. J. J. Am. Soc. Mass Spectrom. 1994, 5, 779-787 and U.S. Pat. No. 5,160,840 to Vestal.
It is an object of the present invention to provide an improved method for focusing the initial ion velocities in a TOF mass spectrometer and, thus, to achieve higher mass resolution of the instrument. Better focusing also allows one to decrease the voltage used for extraction of ions from an ion source that results in smaller size of the instrument.
It is also an object of the present invention to solve the problem of an ideal (or infinite order) velocity focusing in a reflectron TOP mass spectrometer using a curved electric field inside a reflectron.
In the present invention, in addition to the field-free region, the curved field in the reflectron takes into account also acceleration and deceleration fields in upstream (from the ion source down to the reflectron) and downstream (from the reflectron down to the ion detector) regions, which are always present in any TOF-MS. The reflectron includes a decelerating section and a correcting section, with curved electric fields in the correcting and/or decelerating sections of the reflectron being considered. Moreover, analytic expressions are provided for calculating the profiles of the curved electric field in the second (correcting) section of the reflectron, which expressions are valid for arbitrary electric field distributions in the upstream and downstream regions as well as in the first (deceleration) section of the reflectron. These profiles will depend on the electric field distributions in the upstream and downstream regions and in the first (deceleration) section of the reflectron.
The use of the curved field results in ideal (infinite order) focusing, in contrast to the limited (first or second) order focussing that is conventionally used. Additionally, the curved field in the present invention can be designed for any geometry of the upstream and downstream regions and the decelerating section of the reflectron, in contrast to the precise adjustment of the lengths of all regions to achieve focusing as has been previously required.
The present invention also provides an improved apparatus for focusing the initial ion velocities in a reflectron TOF mass spectrometer that consists of a two-stage reflectron with a curved profile for the electric field in the second (correcting) section of the reflectron. The profile of the electric field in the first (decelerating) section of the reflectron can be chosen arbitrarily. The profile in the second section depends on the electric fields in other sections of the TOF mass spectrometer. The two sections of the reflectron can be separated by a grid, or a gridless design can be utilized.